Synthesis and cholesterol lowering abilities of novel prebiotic galacto oligosaccharides produced using lactobacilli reverse enzyme synthesis

• Main Author: Stephens, Katherine
• Published: University of Reading 2017
• Online Access: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.736205

Cardiovascular diseases (CVD) are increasing in the UK and worldwide. Coronary heart disease (CHD) is caused by atherosclerotic plaques which may be formed due to high serum lipid levels. Probiotics have been associated with many health benefits one of which is cholesterol and triglyceride reduction. This study screened twelve Lactobacillus spp. in pure culture for their ability to remove cholesterol, deconjugate bile acids and exhibit high bile salt hydrolase (BSH) activities; important traits for a lipid reducing probiotic. Screening of the candidate probiotics found cholesterol removal of 5-37% after 24 hours. Not all probiotics exhibited high bile salt hydrolase activities, however a preference towards glycholate was observed. This correlated well with deconjugation ability, with all probiotics tested exhibiting significantly higher rates towards glycholate deconjugation. Four probiotics L.fermentum 11976, L.fermentum 30226, L. rhamnosus OR-l and L. plantarum 2830 were selected for further testing and prebiotic generation. Galactooligosaccharides (GOS) are recognised prebiotics which have been shown to exert bifidogenic effects and improve gut health. GOS can be synthesised using ß-galactosidases (ß-gal); enzymes which are found within lactobacilli. The four probiotics were tested for their ability to synthesise GOS. First, analysis of ß-gal was carried out investigating enzyme expression, pH and temperature stability. GOS was then synthesised using whole cell extracts from each of the four probiotics; producing GOS yields of 36-45% at 80% lactose conversion. The GOS mixes were purified using Saccharomyces cerevisae and Kluyveromyces morxianus which produced 90% pure GOS. Trisaccharide composition of the purified GOS's were analysed by gas chromatography flame ionisation detector (GC-FlD), this showed that structures of synthesised GOS differed from one another and the positive control. Finally, the selectivity of the GOS' were analysed in mixed culture using pH controlled batch fermentations. The synbiotic of synthesised GOS with its parent probiotic were included, as well as a positive control, Bimuno (BGOS) and synbiotic of Bimuno plus probiotic. Samples were taken to analyse bacterial populations; short chain fatty acids, cholesterol reduction and BSH activities and deconjugation abilities. All four GOS from lactobacilli significantly increased lactobacilli populations. L. fermentum 11976 GOS did not significantly increase bifidobacteria, this is the first GOS exhibited to selectively enhance lactobacilli populations. All GOS's exhibited cholesterol reducing abilities and deconjugation abilities which were enhanced using the synbiotic of lactobacilli GOS and parent probiotic strain. In conclusion, this study has generated four novel GOS with the ability to increase lactobacilli populations, reduce cholesterol and deconjugate bile acids in vitro. This work shows promise for future in vivo investigations.